The major goal of this Program Project is the development of robust, exportable technology for high throughput DNA sequencing, developed and tested in the context of a medium-scale production sequencing environment. The project builds upon the technology developed in the Smith laboratory at the University of Wisconsin (UW) over the past several years for an integrated system for sequencing comprised of three major parts: a high throughput robotics system for sample preparation, a novel electrophoresis platform for fluorescence-based automated DNA sequencing, and a software system for data analysis. Over 2 million bases of raw data have been produced with this system to data, including the reactions, electrophoresis and fluorescence detection, and the sequence data processing and base-calling. All of the technology is robust, modular, exportable, and available to the community. Average read lengths are >1000 bases. The Program Project is structured as a close collaboration with the chromosome 19 mapping group at Lawrence Livermore National Laboratory, who are responsible for producing contiguous sequence-ready clones covering a region of 10 Mb of human DNA on chromosome 19 and 2 Mb of synthetic mouse DNA. Two major emphases of the technology development efforts are a) increasing throughput of the automated system to a level of 6 Mb/yr, and b) automated major aspects of the finishing process. The throughput of the integrated systems will be increased to a level of 6 Mb/yr by the end of the grant period, with staffing requirements of approximately 6 FTEs to operate at that level.